This research proposal is designed to develop a unique model system to study the interactions of steroid hormones in target cells and the biochemical mechanism of hormone action. Antheridiol is a steroid hormone which induces sexual differentiation of "male" hyphae of the comycete fungus, Achlya ambisexualis. This water mold is the most primitive eukaryotic organism known to contain steroid hormone system and it should offer several advantages for investigations on hormone action. Although some aspects of this interesting system have been studied in detail, there is essentially no information on the fate of antheridiol in target cells or its interaction with receptors. For such a study, a radioactive form of antheridiol with high specific activity is needed and the initial aspect of the present proposal is to prepare this tritium labeled antheridiol. One partially-tested procedure plus several alternative methods of synthesis will be employed. When the radioactive hormone has been prepared, its fate in target cells will be examined. The metabolism and intracellular localization of (3H) antheridiol will be measured at various times after its administration to cell cultures. These results will be compared with the time-course of various hormonal responses such as increased synthesis of RNA and protein and the appearance of antheridial buds. Efforts will then be made to identify binding proteins for (3H) antheridiol. Hormone complexes will be measured after adding (3H) antheridiol to intact cells or to subcellular extracts. The quantity, specificity, and affinity of the hormone-binding components will be measured as an indication of their biological significance in the action of antheridiol. These investigations should provide an indication as to the degree of similarity between this system and those of higher organisms. They will also provide the basis for future studies on the biochemical mechanism of steroid hormone action.